The therapeutic benefits of enhancing T cell function by blockade of coinhibitory receptor PD-1 and one of its ligands, PD-L1, are now well established for multiple malignancies. Targeting the PD-1 pathway for treatment of chronic infectious diseases is also an area of active development. However, whereas PD-1 blockade unleashes beneficial tumor-specific T cell responses during cancer immuno- therapy, increasing pathogen-specific T cell function through PD-1 blockade can either enhance control of infection or drive lethal immunopathology. Given the long duration of standard treatment regimens for tuberculosis and the increasing prevalence of drug-resistant strains, host-directed therapies that prove particularly useful in TB are desirable. Human (Mtb)specific CD4 T cells express PD-1 during active TB, and levels of PD-1 on Mtb-specific T cells decrease after successful TB treatment. Accordingly, PD-1 blockade has been suggested as a host-directed therapy for, but it is not clear whether PD-1 blockade would be beneficial or detrimental in human TB. Our preclinical data in mice suggest that boosting Th1 responses by targeting PD-1 may be detrimental during Mtb infection. We have previously shown that PD-1/ mice are hypersusceptible to Mtb infection, developing large necrotic lesions with high bacterial loads and succumbing faster than even T celldeficient mice. We also have found that the inability of PD-1/ mice to control Mtb infection is due to increased Mtb-specific CD4 T cell responses, as early mortality in PD-1/ mice is prevented by CD4 T cell depletion. Moreover, we have shown that overproduction of IFN-g by CD4 T cells drives early mortality in Mtb-infected PD-1/ mice. Thus, although I IFN-gproducing TH1 cells are required for host resistance to mycobacteria, their enhanced activity in the absence of PD-1 counterintuitively exacerbates TB in mice. However, it is not clear if PD-1 has a similar host-protective role in humans. In this project, we reported two cases of TB in individuals receiving antiPD-1 monoclonal antibodies as cancer treatment. In one patient with nasopharyngeal carcinoma (NPC), PD-1 blockade was followed by the rapid development of disseminated TB that was eventually fatal. In the second patient being treated with antiPD-1 for Merkel cell carcinoma (MCC), pulmonary TB reactivation was much milder and was successfully treated with anti-TB therapy. In this patient that survived, we found that PD-1 blockade was associated with a burst of Mtb-specific Th1 cells in circulation. Collectively, the preclinical mouse model data and clinical case reports indicate that negative regulation of host immune responses are important in limiting detrimental inflammation during tuberculosis. In this annual reporting period, we also examined the molecules that regulate the migration of CD4 T cells into the lungs of Mtb infected mice. We previously showed in mice that CXCR3+ Th1 cells enter the lung parenchyma and suppress M. tuberculosis growth, while CX3CR1+ KLRG1+ Th1 cells accumulate in the lung vasculature and are nonprotective. We, therefore, quantified the contributions of these chemokine receptors to the migration and entry rate of Th1 cells into M. tuberculosis-infected lungs using competitive adoptive transfer migration assays and mathematical modeling. We found that in 8.6hours half of Mtb-specific CD4 T cells migrate from the blood to the lung tissue parenchyma. CXCR3 deficiency decreases the average rate of Th1 cell entry into the lung parenchyma by half, while CX3CR1 deficiency doubles it. KLRG1 blockade has no effect on Th1 cell lung migration. CCR2, CXCR5, and, to a lesser degree, CCR5 and CXCR6 also promote the entry of Th1 cells into the lungs of infected mice. Moreover, blockade of G-protein-coupled receptors with pertussis toxin treatment prior to transfer only partially inhibits T cell migration into the lungs. Thus, the fraction of Th1 cell input into the lungs during M. tuberculosis infection that is regulated by chemokine receptors likely reflects the cumulative effects of multiple chemokine receptors that mostly promote but that can also inhibit entry into the parenchyma.